Three-dimensional deuterium-carbon correlation experiments for high-resolution solid-state MAS NMR spectroscopy of large proteins

Well-resolved ²H-¹³C correlation spectra, reminiscent of ¹H-¹³C correlations, are obtained for perdeuterated ubiquitin and for perdeuterated outer-membrane protein G (OmpG) from E. coli by exploiting the favorable lifetime of ²H double-quantum (DQ) states. Sufficient signal-to-noise was achieved due to the short deuterium T ₁, allowing for high repetition rates and enabling 3D experiments with a ²H-¹³C transfer step in a reasonable time. Well-resolved 3D ²H_DQ-¹³C-¹³C correlations of ubiquitin and OmpG were recorded within 3.5 days each. An essentially complete assignment of ²H_DQα shifts and of a substantial fraction of ²H_DQβ shifts were obtained for ubiquitin. In the case of OmpG, ²H_DQα and ²H_DQβ chemical shifts of a considerable number of threonine, serine and leucine residues were assigned. This approach provides the basis for a general heteronuclear 3D MAS NMR assignment concept utilizing pulse sequences with ²H_DQ-¹³C transfer steps and evolution of deuterium double-quantum chemical shifts.